ADDITIONAL ORGANIZATIONAL FEATURES OF THE MURINE FOLYLPOLYGLUTAMATE SYNTHETASE GENE - 2 REMOTELY SITUATED EXONS ENCODING AN ALTERNATE 5'-END AND PROXIMAL OPEN READING FRAME UNDER THE CONTROL OF A 2ND PROMOTER

Nucleotide sequence analysis of independently isolated clones from a m ouse liver cDNA library identified two additional splice variants of f olylpolyglutamate synthetase (FPGS) mRNA with novel sequence at the 5' end, These variants incorporate two new alternatives (exons Ala and A lb) of exon 1 in the murine FPGS gene which are also spliced to exon 2 , Exon Ala encodes most of the 5'-untranslated region, Exon Alb encode s a downstream segment of the 5'-untranslated region, two ATG start co dons, and a unique mitochondrial leader peptide as well as 15 addition al amino acids of cytosolic FPGS not encoded by all previously identif ied (Roy, K,, Mitsugi, K,, and Sirotnak, F. M, (1996) J, Biol, Chem,, 271, 23820-23827) splice variants, It was also found by direct sequenc ing of genomic fragments that although exon Alb is spliced to exon 2, these new alternatives (i.e. exons Ala and Alb) to exon 1 are found ap proximately 9.5 kilobases upstream from exons Bla, Bib, and B1c, Exons Ala and Alb are separated from each other by a 124-nucleotide intron, Sequencing of the region 5' to exon Ala revealed a nucleotide sequenc e that was promoter-like and different from the downstream promoter re gion in the content of putative cis-acting elements, Primer extension analysis identified a number of potential transcription start sites wi thin the more 3' end of this region, FPGS RNA transcripts incorporatin g exons Ala and Alb were detected in both normal mouse tissues, partic ularly, liver and kidney, and also to a varying extent in tumors; FPGS RNA transcripts incorporating exons Bla, Bib, and B1c were detected m ainly in tumors, Thus, transcription of the FPGS gene in this tissue-s pecific manner appears to reflect the different usage of alternates to exon 1 under the control of different promoters, An unusual splice va riant identified infrequently in a mouse liver cDNA library was 2.67 k ilobases in size and incorporated exons Ala and Alb and a segment of t he downstream promoter region along with exons B1c and B1b and exons 2 -15.